1. Field of the Invention
This invention relates to a transducible composite plasmid and more particularly to a composite plasmid capable of transduction having a Coryneform glutamicacid-producing bacterium as the host thereof.
2. Description of the Prior Art
Some species of the Coryneform glutamic-acid-producing bacteria are known to produce large amounts of L-glutamic acid, and some other species, particularly certain mutants of wild strains, produce large quantities of amino acids such as lysine and purine nucleotides such as inosinic acid. These large producers are industrially important microorganisms. Recently, attempts have been made to commercially breed and improve microorganisms using the recombinant DNA techniques. Development of recombinent DNA techniques is also now well under way with respect to Coryneform glutamic acid-producing bacteria. For example, the development of a host-vector system for Corynebacterium glutamicum, the development of a host-vector system for Brevibacterium lactofermentum, and the breeding of threonine-producing microorganisms are reported respectively in Glossary of Lectures at the 1983 General Meeting of Japan Agricultural Chemical Society, page 333 (1983) and Glossary of Lectures at the 1983 General Meeting of Japan Fermentation Engineering Society, page 283 and page 284 (1983). Composite plasmids disclosed in European Application No. 83302478.9 (publication no. 0093611; U.S. application Ser. No. 386,980) are also advantageous vectors for use in Coryneform glutamic-acid-producing bacteria as the hosts in recombinant DNA investigations.
The vector systems heretofore known for host Coryneform glutamic-acid-producing bacteria invariably use plasmids as their vectors. For successful insertion of these plasmid vectors into their hosts, separation of the plasmid vectors and subsequent treatment of separated plasmid vectors (by such complicated means of transformation as the protoplast method or the calcium treatment method) are indispensable.
In Escherichia coli, there is available the socalled cosmid, i.e., a recombined plasmid possessing a DNA region including a cohesive end (COS) originating in the lambda phage [Fukumaki, U., Shimada, I., & Takagi, Y., Proc. Natl. Acad. Sci. USA, 73, 3238 (1976) and Collins, J. Hohn, B.: Proc. Natl. Acad. Sci. USA, 75, 4242 (1978)].
This cosmid is characterized by its ability to be easily transmitted from a host strain to another strain by a transduction method using a phage. Herein, the term "transduction method" is defined as the insertion of a gene through the medium of a phage particle, as described in Protein Nucleic Acid-Enzyme, extra issue, "Experimental Methods for Microorganism and Phage Heredity," page 64 (1972). This method is based on the principle that when a cosmid-retaining strain of bacteria is infected with an intact phage, for example, there is formed a pseudo-phage particle containing cosmid DNA instead of natural intact phage DNA in a fixed proportion, and this particle acquires the ability to effect introduction of the cosmid DNA into a new host strain. When this cosmid is adopted, the desired transmission of genetic information is effected by simply preparing the phage bacteriolyzate of a donor bacterium and mixing this bacteriolyzate with a recipient bacterium, thereby causing infection. Thus, this method accomplishes the transmission of genetic information very rapidly and conveniently as compared to other methods using a plasmid.
In none of the Coryneform glutamic-acid-producing bacteria, however, is there found a vector capable of transduction. Thus, none of the known vectors permit transmissible of a plasmid by the transduction method.